A number of methods are already known for treating membranes of biological cells and/or their cell components by way of exchanging biological material between them. In recent years various methods of transmitting biological material through the membrane of a cell have increasingly gained in importance. In this method, membrane-impermeable molecules are sluiced through pores which have been formed in the membrane through external forces.
It is known to treat cell membranes with chemical substances to make them permeable. In particular these include pore-forming and/or diffusion-promoting compounds. In this context peptide antibiotics, such as valinomycin, and detergents such as sodium dodecylsulphate or triton X-100 can be used.
Another possibility of making the membranes of biological cells permeable is electrical permeabilisation. This includes methods such as electrotransfection, electroporation and electrofusion. These methods are carried out in macroscopic devices as well as in Microsystems or microstructures.
Electrical permeabilisation has the advantage that it can be implemented without a vehicle and without externally added chemicals. Electropermeabilisation by means of an electrical field, also known as “electroporation”, has for some time been an established method of taking up free DNA in, for example, eukaryotic cells. This is done by exposing the eukaryotic cells to an electrical field with high field strength in the presence of DNA. It is presumed that as a result of the “electric shock” pores are temporarily formed in the membrane. This allows the DNA to be transmitted and to flow into a biological cell (Zimmermann and Neil, 1996 [Zimmerman, U. and Neil G. A. (Eds))[1996] Electromanipulation of Cells, CRC Press, Boca Raton).
In practice it has been found that often not necessarily all of the biological cells are influenced by the electrical field in the same way. In particular in the case of the fusion of smaller and larger cells the problem arises that in general the large cells are destroyed as the critical field strength for these cells is less than for small ones (integrated Laplace equation) (Zimmermann and Neil; citation see above). During the destruction of the cells, i.e. when the membrane bursts, proteolytic enzymes are released which can modify the membrane proteins of the intact and/or fused cells. This is associated with considerable drawbacks. The released proteolytic enzymes affect the fused cells products to the extent that they cannot carry out the intended function. This becomes a major problem especially if, for example, allogenic dendritic cells have been treated in the electrical field and the resulting products of fusion have been so modified in their membrane structure that they do no produce the desired immune response after injection into the patient. Similar problems can occur during the production of hybridoma cells through the electrofusion of B-lymphocyte cells with myeloma cells. Thus it has been observed that due to the destruction of cells in the electrical field, proteolytic cells are present in the medium which negatively affect the hybridome clones produced during electrofusion.